Dopamine-B-hydroxylase (DBH, EC 1.14.17.1) is a copper containing enzyme that catalyzes the synthesis of the neurotransmitter, norepinephrine, in the sympathetic nervous system. It is the object of this proposal to study in detail the mechanism of inhibition of dopamine-B-hydroxylase by certain compounds, some of which have been used to treat hypertension in man or to manipulate the endogenous levels of the neurotransmitter, norepinephrine, in experimental animals. Quantitative study of the interaction of these inhibitory species with DBH using both spectroscopic techniques such as electron paramagnetic resonance (epr), UV, VIS, NIR electronic absorption, fluorescence emission, and nuclear magnetic resonance (nmr) and chemical methods will also yield estimates of the number and copper stoichiometry of the active sites in DBH, neither of which are currently known. Chemical and spectroscopic studies of the native and apo enzyme will be used to elucidate the nature of the ligands and coordination geometry of the essential copper in DBH. The way in which molecular oxygen is activated and utilized by DBH will also be investigated. These studies will elucidate the mechanisms involved in the short term regulation of DBH and norepinephrine biosynthesis.